Phosphorylation events associated with different states of activation of a hepatic cardiolipin/protease-activated protein kinase. Structural identity to the protein kinase N-type protein kinases.
Abstract
Cardiolipin- or protease-activated protein kinase, isolated from rat liver cytosol and originally named liver PAK-1, was found to be the natural form of protein kinase N (PKN) by comparing the sequences of 43 tryptic peptides of the purified liver enzyme and determining the corresponding liver cDNA sequence. These analyses also identified (i) Arg-546 as the major site of proteolytic activation, (ii) the protease resistance of the C-terminal extension beyond the catalytic domain, and (iii) in vivo stoichiometric phosphorylation of Thr-778 in the mature enzyme. Homology modeling of the catalytic domain indicated that phosphothreonine 778 functions as an anchoring site similar to Thr-197 in cAMP-dependent protein kinase, which stabilizes an active site compatible with preferred substrate sequences of PAK-1/PKN. Sigmoidal autophosphorylation kinetics and increased S6-(229-239) peptide kinase activity following preincubation with ATP suggested phosphorylation-dependent activation of PAK-1/PKN. The onset of activation corresponded with phosphorylation of the regulatory domain site Ser-377 (located within a spectrin homology region), followed by Thr-504 (within a limited protein kinase C homology region), and, to a lesser extent, Thr-...Continue Reading
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Citations
1H, 15N and 13C resonance assignments of the HR1c domain of PRK1, a protein kinase C-related kinase.
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